Cell or drug encapsulation device having a wet seal

ABSTRACT

A biologically implantable containment device having a wet seal, the device being adaptable for drug formulations or cell suspensions. A porous membrane, in a tubular configuration, is formed and can be configured as part of a closed cell-tight system for loading. During loading, the containment device membrane is wet, while the loading system remains cell-tight. The containment device is wet-sealed through a combination of heat and pressure, while the system remains cell-tight. Sealing the containment device substantially or completely eliminates metabolic functioning of any organisms in the vicinity of the closure. The wet-seal is formed by melting a thermoplastic material that is in contact with the membrane. The containment device is separated from the cell-tight loading system, which remains closed after separation.

CROSS REFERENCE TO RELATED APPLICATIONS

[0001] This application is a division of co-pending application Ser. No.09/515,264 filed Feb. 29, 2000.

BACKGROUND OF THE INVENTION

[0002] 1. Field of the Invention

[0003] The invention relates to the field of implantable devices andmore particularly to devices that are wet-sealed.

[0004] 2. Background Information

[0005] Within the field of implantable devices, it is known to providepermeable membrane structures for implantation, the structuresconfigured to hold drug formulations or cellular suspensions. A numberof techniques have been proposed to form those structures and seal thestructures. In the majority of those known techniques, the device ismanufactured without the cellular suspension or drug formulation.Subsequent loading of the cellular suspension or drug formulation mayoccur outside a host or after the device is implanted into the host.When a suitable cell suspension or drug formulation is loaded into thedevice, it is typical and frequently desirable for the permeablemembrane to become wet with fluid. Given the nature of the membranes, itis known that sealing a wet membrane can be difficult or impossible.This is because known glues and solvents that are appropriate formembranes in a dry state are frequently not compatible with a wetmembrane, or are toxic to cell suspensions loaded into the membranestructure. To offset this difficulty, different dry and wet sealtechniques have been proposed.

[0006] In one technique, such as disclosed in U.S. Pat. No. 5,902,745 toButler et al., the device includes a permeable tubular membrane, whichis sealed with a mechanical seal after loading the device with anappropriate cell suspension. In this technique, the membrane is wet whenthe seal is formed, but seal integrity relies on the quality of themechanical seal. With implantable devices, the mechanical sealdimensions are small and can be difficult to reliably manipulate. Inaddition, because the loading and sealing operations can be distinct,there is an opportunity for contamination of the device exterior withcells from the cell suspension after the loading operation.

[0007] In another technique, such as disclosed in U.S. Pat. Nos.5,653,687; 5,653,688; 5,713,887; 5,738,673 and 5,932,460 issued to Millset al, a dry seal is formed after the device is loaded. However, theloading and sealing steps are distinct and the device is open to theloading environment after loading and before the device is sealed. Forsome of these seals, the seal depends on mechanical aspects of the seal.Some of the disclosed seal techniques require a solvent based seal. Thesolvents described may be toxic to the cell suspension, however. In oneparticular embodiment of the seal, a portion of the device is broken offand removed after loading and prior to sealing. This action presents astrong possibility of contaminating the loading environment. Thiscontamination can be subsequently transferred to the exterior of thedevice, or to other devices or apparatus.

[0008] In another technique, such as disclosed in U.S. Pat. Nos.5,545,223 and 5,549,675 issued to Neuenfeldt et al., the apparatus ordevice is first implanted in a host and then loaded with a cellularsuspension in the host environment. In addition to problems that aredescribed with wet sealing of the device, this technique is performedthrough an incision or injection port following implantation in thehost, thereby exposing the device and the host to a risk ofcontamination. The technique of Neuenfeldt et al. also requires a largerdevice to accommodate the distance between the cell suspension and theseal. This larger device also produces greater host trauma duringimplantation. In some of the known techniques, the device or apparatusis loaded in an area that is remote from the host. In these methods, theloading process or apparatus provides opportunities for contaminationfrom drug formulations or cell suspensions between the loading and thesealing steps.

[0009] As described, the methods available do not provide a secure andreliable closure system, that reduces the possibility of contaminationduring loading. In addition, the methods available do not provide amethod to reliably seal a device after the membrane is wet. Systems andmethods to address these and other deficiencies are needed.

SUMMARY OF THE INVENTION

[0010] In one aspect, the present invention provides a method of closinga containment device that comprises wetting at least a portion of apermeable polymeric membrane of the containment device with a liquid andapplying heat to at least a portion of a wetted thermoplastic polymer inassociation with the membrane to create a closure. Such a closure isreferred to herein as a “wet seal.” In this “wet sealing” process, thethermoplastic polymer melts at a lower temperature than the polymericmembrane. Once melted, the thermoplastic polymer integrates with thepolymeric membrane and flows along surfaces and into availableinterstices of the membrane. Through passageways become filled with themelted polymer, thereby blocking fluid communication in the polymericmembrane in the region of the closure. When the thermoplastic polymercools below its melt temperature, a closure is formed in the device. Theclosure is cell-tight and often liquid-tight. The portion of the devicehaving a closure formed with a wet seal delineates a cell-impermeableregion of the device. The application of heat may be accompanied byslight pressure and a heat sink may be applied to limit heat transferbeyond the closure region to the permeable membrane. After forming theclosure, the method may include pressure checking the closure integrity.The device may include additional closures that are formed by wet or drysealing techniques.

[0011] In one aspect, the present invention provides a method of closinga containment device that comprises wetting a porous expandedpolytetrafluoroethylene (ePTFE) membrane of the containment device witha liquid, and applying heat to a portion of the membrane incommunication with a thermoplastic polymer, such as fluorinated ethylenepropylene (FEP), to create a closure. The closure is formed by meltingand fusing of the polymer to itself and the membrane in the presence ofthe liquid.

[0012] In one aspect, the present invention provides a method of closinga containment device comprising wetting a permeable membrane of thecontainment device with a liquid, wetting a thermoplastic polymer regionof the device with the liquid and applying heat directly to thethermoplastic polymer region to create a closure. In this aspect, thethermoplastic polymer region is joined to the permeable membrane beforewet sealing the containment device.

[0013] In one aspect, the present invention provides a method of closinga containment device that comprises applying sufficient heat to aportion of a permeable membrane in association with a thermoplasticpolymer to melt and flow the thermoplastic polymer, followed by twistingthe membrane/thermoplastic polymer combination in the region of theheating to form a closure. The membrane/thermoplastic polymercombination is also elongated while heating or twisting the materials.After heating, twisting, and elongation a separation region is formedand the membrane is cut in the separation region. In one aspect, thepresent invention provides a containment device comprising a membrane, apolymer in communication with the membrane, and a closure. The closureis created by applying heat to a portion of the membrane and a portionof the polymer after wetting the membrane with a liquid.

[0014] In one aspect, the present invention provides a containmentdevice comprising a membrane, a polymer region joined to the membraneand a closure. The closure is created by applying heat directly to thepolymer region after wetting the membrane and the polymer region with aliquid.

[0015] In one aspect, the present invention provides a containmentdevice comprising a membrane and a closure. The closure is created byapplying heat to a portion of the membrane and twisting the membrane inthe region of the heating. In one aspect, the present invention providesa method of forming a containment device. The method comprises forming acontainment region that includes a membrane, forming a thermoplasticpolymer region joined to the membrane and forming a closure region. Theclosure region communicates with the containment region and applyingheat directly to the thermoplastic polymer region after wetting themembrane creates a closure in the closure region.

[0016] In one aspect, the present invention provides a method of forminga containment device. The method comprises forming a containment regionthat includes a membrane, and forming a closure region. The closureregion communicates with the containment region and applying heat to aportion of the membrane and twisting the membrane in the region of theheating creates a closure in the closure region.

[0017] In one aspect, the present invention provides a method of loadinga containment device comprising placing a cell suspension or drugformulation in a containment region of the device through a closEableopening, the containment region including a membrane and the liquidwetting the membrane, and creating a closure in the closeable opening byapplying heat to a portion of the membrane in association with athermoplastic polymer.

[0018] In one aspect, the present invention provides a method of loadinga containment device comprising creating a closed cell-tight system, thesystem including the containment device and a source of metabolicallyactive cells. Loading a containment region of the device with themetabolically active cells via a closure region, the containment regionincluding a membrane. Creating a closure at the closure region, theclosure substantially or completely eliminating metabolically activecells in the vicinity of the closure, and subsequent separation of thesource of cells while maintaining a closed cell-tight system.

[0019] The foregoing specific aspects, objects and advantages of theinvention are illustrative of those which can be achieved by the presentinvention and are not intended to be exhaustive or limiting of thepossible advantages that can be realized. Thus, the aspects, objects andadvantages of this invention will be apparent from the descriptionherein or can be learned from practicing the invention, both as embodiedherein or as modified in view of any variations which may be apparent tothose skilled in the art. Accordingly the present invention resides inthe novel parts, constructions, arrangements, combinations andimprovements herein shown and described.

BRIEF DESCRIPTION OF THE DRAWINGS

[0020] The foregoing features and other aspects of the invention areexplained in the following description taken in conjunction with theaccompanying figures wherein:

[0021]FIG. 1 illustrates a containment device of the instant inventionand an implantable containment apparatus suitable for holding thecontainment device;

[0022]FIG. 2 illustrates a cross-section of an embodiment of thecontainment device of the instant invention;

[0023]FIG. 3 illustrates an embodiment to form the tubular membrane ofthe instant invention; FIG. 4 illustrates various cross-sectionembodiments of the containment device of the instant invention;

[0024]FIG. 5 illustrates an embodiment of the containment device of theinstant invention;

[0025]FIG. 6 illustrates an embodiment of a dry seal on an end of thecontainment device of the instant invention;

[0026]FIG. 7 illustrates a cross-section of an embodiment of thecontainment device of the instant invention;

[0027]FIGS. 7A-7C illustrate cross-sections of embodiments of thecontainment device of the instant invention;

[0028]FIG. 8 illustrates an embodiment of a heat source used to formclosure regions or seals in containment devices of the instantinvention;

[0029]FIG. 9 illustrates an embodiment of steps to form a containmentdevice of the instant invention;

[0030]FIG. 10 illustrates a loading hood used in one embodiment forloading the containment device of the instant invention;

[0031]FIG. 11 illustrates a loading jig in one embodiment for loadingthe containment device of the instant invention;

[0032]FIG. 12 illustrates an embodiment of a loading apparatus attachedto a containment device of the instant invention;

[0033]FIG. 13 illustrates an embodiment of loading cell suspension intoa loading apparatus attached to a containment device of the instantinvention;

[0034]FIG. 14 illustrates an embodiment of loading cell suspension intoa containment device of the instant invention and clamping thecontainment device after the cell suspension is loaded;

[0035]FIG. 15 illustrates an embodiment of heat sealing or closing acontainment device of the instant invention using an electrically heatedclamp;

[0036]FIG. 16 illustrates an embodiment of twisting and elongating theclosure region of the containment device of the instant invention;

[0037]FIG. 17 illustrates an embodiment of separating the sealedcontainment device of the instant invention from the loading apparatus,and the resulting closure at the closure region;

[0038]FIG. 18 illustrates an embodiment of steps to close or seal acontainment device of the instant invention;

[0039]FIG. 19 illustrates alternative embodiments of the containmentdevice of the instant invention;

[0040]FIG. 20 illustrates an alternative embodiment of the containmentdevice of the instant invention; and

[0041]FIG. 21A illustrates an alternative embodiment of the containmentdevice of the instant invention.

[0042]FIG. 22 is a graph showing the presence of chemical groups fromresidue associated with wet seals of the instant invention.

[0043] It is understood that the drawings are for illustration only andare not limiting. It is specifically understood that the scale ofelements and relative dimensions in the drawings may be exaggerated toprovide clarity and illustrate individual aspects.

DETAILED DESCRIPTION OF THE INVENTION

[0044] The invention provides a containment device, methods of makingthe device and methods of loading and sealing the device. Thecontainment device is particularly suited for use as a medical device,such as a cell encapsulation device, a drug delivery device, or a genetherapy device. The containment device may be inserted into a previouslyimplanted containment apparatus residing within a recipient, such as ananimal or human, or it may be implanted directly in a recipient. Thedevice includes a permeable membrane, which partially defines anenclosed space of the device, and a closure region of the device.Materials (e.g., cells, or drugs) are loaded into the device from aloading device through the closure region into a containment region,after which the closure region is treated to form a closure. The closureis typically created by heating a thermoplastic polymer associated withthe permeable membrane in the presence of a liquid that wets at least aportion of the membrane in the closure region. The liquid may also wetat least a portion of the thermoplastic polymer. Wetting of the membraneis often a result of filling the device with a liquid, such as a cellsuspension or drug formulation, or from sterilization procedure prior tofilling. Forming the closure may include pressure or clamping as part ofthe sealing process. A closure formed in this manner is referred toherein as a “wet seal.”

[0045] Different embodiments for creating the closure after loading thecontainment device are disclosed. In one embodiment, the closure iscreated when heat is applied to a portion of a tubular porous expandedpolytetrafluoroethylene (ePTFE) membrane, which includes a tube offluorinated ethylene propylene (FEP) thermoplastic polymer inassociation with the ePTFE membrane. The applied heat is sufficient tomelt the FEP thermoplastic, but not of a magnitude to melt orsubstantially degrade the ePTFE membrane. While the FEP is in a meltedstate, the tubular membrane in the region of the melted FEP is pressedtogether, preferably by twisting the tubular device around itslongitudinal axis, and elongated, thereby creating the closure. Ineffect, the FEP and ePTFE form a fused or welded cell-free andcell-tight wet seal closure. After forming the wet seal closure, thecontainment device is separated from the cell loading device by a cutthrough the fused ePTFE/FEP material in the closure region.

[0046] After a wet seal is formed, thermally degraded residue from cellsor cell culture media can usually be found on surfaces of material inthe closure region. Residue may also be embedded in material of theclosure region. The presence of such residue can be detected by samplingcharred material from the closure region and subjecting the sample and acontrol to analysis. A preferred analytical method is Fourier TransformInfrared Spectroscopy (FTIR). In comparison with a control, chemicalgroups indicating degradation of cells or cell culture media in the wetseal region become evident as changes in peaks on a graph. An example ofsuch a graph is shown in FIG. 22. In the graph, the curve represented bya dotted line is from a control sample. The curve represented by acontinuous line are peaks generated from a test sample. Differences inthe peaks on the graph are indicative of chemical changes in the cellculture media used to load cells into a device of the present inventionat the time of a thermally effective wet sealing process in the closureregion. Such residue is only present in the closure region if a wet sealhas been formed in the region.

[0047] Another important aspect of the invention is that the containmentdevice and the loading device form a closed cell-tight system duringloading, sealing and separation. This closed cell-tight aspect of thesystem is maintained during the loading and closure or sealing andsubsequent separation of the containment device. In this manner,contamination of the device exterior by cells from either the loadeddevice or the cell delivery device is completely or substantiallyprevented.

[0048] During trials, containment devices were manufactured, loaded,sealed and later implanted according to the instant invention. Thesetests positively demonstrate that the methods and apparatus disclosed inthe instant application accomplish the stated objectives of reducing oreliminating contamination of the device exterior and containing cellswithin the device. In vitro testing demonstrated that exteriorcontamination during device loading was reduced or eliminated by usingthe disclosed method and system. In vivo testing for up to 6 monthsshowed that cell containment within the containment device was possiblewithout device or seal failure. These test results provide strongevidence that containment devices, manufactured, loaded, sealed andimplanted according to the instant invention, will maintain the desiredisolation of cells within the device.

[0049] Another important aspect of the invention is that forming theclosure through clamping or pressure does not compromise the mechanicalintegrity of the containment device.

[0050] Uses or Applications of the Device

[0051] The containment device according to the instant invention can bedirectly implanted in a recipient in need of treatments provided by thecontents of the device. When directly implanted in a recipient, knownsurgical techniques are used to prepare an implantation site andposition the containment device in the recipient. This includes anincision at the site and preparation of a tissue envelope to hold thecontainment device. For ease of insertion, incisions may be used toallow threading of the containment device between the incisions. Theimplantation site can be a subcutaneous location that is somewhatprotected from external forces and not subject to significant flexduring normal recipient activities. As examples, the inner forearm orthe inner upper thigh of a human may be appropriate sites.

[0052] The containment device can also be indirectly implanted in arecipient with the used of a containment apparatus for the device. Suchan apparatus is disclosed in U.S. Pat. No. 5,843,069, issued to Butleret al. (the '069 patent) and incorporated herein by reference. Referringto FIG. 1, the containment device 100 according to the instant inventioncan also be placed or replaced in an implantable containment apparatus120 of the '069 patent. The apparatus and methods disclosed in the '069patent overcome some of the above-described problems with directimplantation. When the containment device 100 of the instant inventionis used with the implantable containment apparatus 120 of the '069patent, the containment device is first prepared or filled with theappropriate cell suspension or drug formulation. After prepared orfilled, the containment device is wet sealed, as described in greaterdetail below. Once wet sealed, the containment device is placed orreplaced in the implantable containment device of the '069 patent.

[0053] In a preferred embodiment, the containment device provides asealed environment for cellular suspensions or drug formulations.However, the device has other embodiments and applications where apermeable membrane encloses a space and the device is placed within ahost or recipient. For example, as micro-machining techniques becomemore common, it will be appropriate for the containment device 100 tocontain a micro-miniature factory, the factory providing different formsof processing. In this embodiment, the factory performs any number ofdifferent manufacturing functions. These include neutralizing orchanging the composition of molecules, substances or compounds in therecipient. The micro-miniature factory within the containment device ofthe instant invention can also produce drugs or compounds that areharvested from the host or recipient. The factory is passive, such as acatalyst, or it is active, with a power source such as internalmetabolism of proteins from the nutrient, or external power from outsidethe host or recipient. In short, the invention does not envisionlimiting the materials within the containment device to only cellsuspensions or drug formulations.

[0054] Materials for the Membrane of the Present Invention

[0055] The containment device of the instant invention includes apermeable membrane. Preferred permeable membranes are much like thepermeable membrane of the '069 patent's implantable containmentapparatus. The membrane of the instant invention allows transport ofnutrients, cellular wastes, and other materials through and across themembrane, but the membrane prevents cell movement or migration throughand across the membrane.

[0056] Referring to FIG. 2, the exterior tube 202 of containment device100 of the present invention is made, primarily, of a permeablepolymeric material having sieving properties. The sieving properties ofsuch permeable polymeric materials can be adjusted to control passage ofsolutes, biochemical substances, viruses, or cells, for example, throughthe material, primarily on the basis of size. Preferred permeablepolymeric materials in the present invention are porous. In general, asthe average pore size of a porous polymeric material increases,increasingly larger biochemicals and biological entities are able topass through the material. In the present invention, porous polymericmaterials capable of preventing the passage of biological cells throughthe material, while permitting biological molecules to pass through thematerial, are most preferred.

[0057] Porous polymeric materials suitable for construction of theexterior tube 202 of a containment device of the present inventioninclude, but are not limited to, porous expanded polytetrafluoroethylene(ePTFE); porous polypropylenes (PP), such as Celgard® (CelaneseSeparations, Inc., Charlotte, N.C.), Solvex® (Millipore Corporation,Bedford, Mass.), and Metricel® (Gelman Sciences, Ann Arbor, Mich.);porous polyethylene (PE), including stretched and sintered forms; porouspolyvinylidene fluoride or PVDF (e.g., Durapore®, Millipore Corporation,Bedford, Mass., or FP Vericel®, Gelman Sciences, Ann Arbor, Mich.);track-etched and other porous polycarbonates (e.g., Isopore®, MilliporeCorporation, Bedford, Mass.); woven or non-woven collections of fibersor yarns, or fibrous matrices, such as those described by Fournier etal. in U.S. Pat. No. 5,387,237 (incorporated herein by reference); orfoams of polyvinyl alcohol (PVA), polypropylene (PP), or polyethylene(PE), either alone or in combination.

[0058] Other materials suitable for construction of exterior tube 202include polymers such as biocompatible polyamides (FH-66®, Gambro AB,Lund, Sweden); cellulosics such as cellulose acetate, nitrocellulose,and mixtures thereof (e.g., NC®, Schleicher and Schuell, Inc., Keene,N.H., and MF®), Millipore Corporation, Bedford, Mass., or Metricel®,Gelman Sciences, Ann Arbor, Mich.); polyacrylamide and its copolymerswith acrylic acid and acrylonitrile (e.g., Hypan®, Hymedix, Inc.,Dayton, N.J.); polyacrylonitrile or PAN and its copolymers, includingwith sodium methallysulfonate (AN-69®, Hospal, Lyon, France) and poly(acrylonitrile-co-vinyl chloride) or (PAN-PVC); porous poly(ether etherketones) or PEEKs; porous polysulfones including poly (ether sulfones)or PESs (e.g., Tuffryn® or Supor®, Gelman Sciences, Ann Arbor, Mich.);or stable, strong, biocompatible hydrogels used in soft contact lenses,such as poly (2-hydroxyethyl methacrylate) or polyHEMA,poly(N-vinyl-2-pyrrolidone) or PVP, and mixtures and copolymers thereof.

[0059] Expanded, porous polytetrafluoroethylene is preferred forconstruction of tube 202. Porous expanded polytetrafluoroethylene(ePTFE) is characterized as a material having void spaces defined bynodes and fibrils. Methods for making ePTFE are taught by Gore in U.S.Pat. Nos. 3,953,566 and 4,187,390, each of which is incorporated hereinby reference. However, the methods for making ePTFE are not subjects ofthe present invention.

[0060] For ePTFE, or similar fibrillated material, the pore size isrelated to the fibril length and fibril density of the material and thethickness of the material. In the present invention, appropriate ePTFEmaterials are selected that resist cellular movement across thethickness of the material, while being selectively permeable tomacromolecules. These materials have microstructures (i.e., fibrillength and fibril density) in combination with material thicknesscontrol in large part the sieving properties, or permeability, of themembrane material. Another approach to characterize the sievingproperties of a porous materials, such as ePTFE, is to measureresistance to fluid flow across the materials. One appropriate measureis the pressure at which a gas can pass through the porous material andform bubbles when the material is immersed in an appropriate liquid.This measurement technique is referred to as a “bubble point” metric.

[0061] Method of Making the Device

[0062] Referring to FIG. 2, in a preferred embodiment, a containmentdevice 100 of the present invention is in the form of a tube. The tubewalls create a central lumen 206, which can hold a drug formulation,cell suspension, or other agent. In a preferred embodiment with a cellsuspension, lumen 206 surrounds a central core 204, which can have anumber of cross-sectional shapes. Examples of the variouscross-sectional shapes are illustrated in FIG. 4, with a star shape 401,a perforated cylinder 403, a randomly porous matrix 405 and a bulbextrusion 407.

[0063] The tube walls 202 are made of a membrane, which comprises anePTFE material. The membrane is a layer of ePTFE material that is a verythin, very strong non-woven web composed substantially of tightly spacedfibrils in which there are essentially no nodes. The fibril lengths haveaverage dimensions ranging between about 0.2 and about 4.0 microns, asmeasured by photomicrography. The fibril density is observed to be veryhigh and tightly packed with multiple points of contact. The points ofcontact do not have sufficient polytetrafluoroethylene material to bereferred to as nodes. The thickness of the material in its finished formis between about 1 micron and about 25 microns. The preferred method ofmaking the membrane utilizes a portion of a method taught by Bacino inU.S. Pat. No. 5,476,589 entitled “Porous PTFE Film And A ManufacturingMethod Therefor,” which is incorporated herein by reference.

[0064] Referring to FIG. 3, a preferred method to form a containmentdevice of the present invention in a tubular shape is by wrapping theePTFE membrane 301, made in accordance with the teachings of Bacino, ona mandrel 303. Longitudinal and helical orientations of the wrapped filmmay be used. A single layer or multiple layers of wrapped film may beemployed. The wraps may be set so as to overlap, or to gap, depending onthe setting of such variables as film width, wrap angle, and mandreldiameter. In many applications, the overlap is preferably about 50%.This construction is then heated from about 340° C. to about 400° C.,preferably 385° C., for about 5-10 minutes to bond the respective layersto each other.

[0065] Another way to form an ePTFE containment device of the presentinvention is through direction extrusion and expansion of the membraneas a tube. However, the various methods to form a tubular membrane arenot key aspects of the invention. A tubular ePTFE membrane, formed asdescribed above, is a hydrophobic membrane. Accordingly, the membranedoes not readily permit liquid water to enter and traverse the void orporous spaces and passages of the membrane. It is known to apply certainalcohols, low surface tension liquids, wetting agents or surfactants tothe ePTFE to render the membrane wettable with liquid water. Thesemethods are described in U.S. Pat. No. 5,902,745 to Butler et al., thedisclosure of which is incorporated herein by reference. However,methods to form a wettable membrane are not subjects of the presentinvention.

[0066] In a preferred embodiment, the membrane is treated to make itwettable. In this embodiment a very dilute aqueous solution of a wettingagent, such as polyvinyl alcohol (PVA) is used. For example, 0.001%polyvinyl alcohol in saline (weight to volume or w/v) provides enoughwetting agent to the ePTFE material to prevent, or limit, spontaneousdewetting of the material. This also prevents, or limits, evolution ofair bubbles in a containment device loaded with cells. The PVA alsorenders the normally opaque ePTFE material essentially translucent totransparent upon wetting. Suitable wetting agents and/or surfactants foruse in this method include, but are not limited to, polyvinyl alcohol,polyethylene glycol, sodium dodecyl sulfate, fluorosurfactants,pluronics, and bile salts in percentages ranging from about 0.001-5.0%.Suitable solvents for this method include, but are not limited to,saline, water, and aqueous buffers, for example.

[0067] In the preferred embodiment of the present invention, wettingagents and/or surfactants are adsorbed onto the surfaces and into thevoid spaces, pores, or passages of the ePTFE membrane and preferablyimmobilized in situ in order to make the ePTFE material wettable withliquid water. There are many ways to immobilize wetting agents orsurfactants, such as, cross-linking, substrate grafting, plasmaimmobilization, ionic complexation, and free radical grafting, etc. Inone example, cross-linking the adsorbed wetting agent or surfactant onthe ePTFE in situ immobilizes the wetting agent or surfactant on theePTFE material. Certain wetting agents or surfactants can be used thatrender ePTFE spontaneously and substantially completely liquid waterwettable. A spontaneously and substantially completely water wettableePTFE material permits liquid water to flow along the surface andthrough the passages of the material by merely contacting the materialwith liquid water. Suitable wetting agents or surfactants for use in thepresent invention include, but are not limited, to polyvinyl alcohol,poly(tetrafluoroethylene-co-vinyl alcohol), polyacrylic acid,polyethylenimine, and polyethylene glycol. Wetting agents and/orsurfactants are adsorbed in various ways, such as solution, or neat,adsorption, vapor deposition, plasma immobilization, and thin filmassembly, for example. Preferably, polyvinyl alcohol is adsorbed toePTFE by adsorbing the polyvinyl alcohol onto the surfaces and into theporous, or void, spaces of the material, followed by immobilization viacross-linking the polyvinyl alcohol to itself with a dialdehyde such asglutaraldehyde.

[0068] A membrane of the present invention made of water wettable ePTFEis strong enough to withstand hydrostatic pressures sufficient to causewater to be forced through the pores of the material across thethickness of the membrane. When water is being forced across thethickness of the membrane, the water wettable ePTFE material functionsas a filter, or an ultrafilter, depending on the permeability of thematerial. As water moves, or seeps, across the thickness of themembrane, it tends to collect into droplets on the outer surface of themembrane. As adjacent droplets grow in size, they merge and run off ofthe cover. This process is referred to herein as “weeping.” Most waterwettable membranes of the present invention are sufficiently permeableto water for pressurized water to visibly weep from the membrane withoutgross channeling of water.

[0069] Ideally, the membrane of the present invention is sufficientlywater permeable to allow the ready separation of aqueous fluid fromcells under relatively low pressure. A ready weep flow of ranging fromabout 0.01 ml/cm²/minute to about 100 ml/cm²/minute at a pressureranging from less than about 3.4*10⁴ Pa to about 6.9*10⁵ Pa shouldpermit relatively rapid cell concentration within the device.

[0070] This is an extremely beneficial attribute of the presentinvention. Unlike other cell containment devices that require cellconcentration before insertion of the cells in the cell device and thencarefully calculated and controlled transfer, the present inventionallows cells to be easily transferred to any desired concentration withminimal pre-concentration steps. Further, by flushing a cell-filledapparatus after initial loading of the cells, a user can assure that allcells are flushed into the device and not left as a wasted residue onthe apparatus.

[0071] Another benefit of the membrane becoming essentially translucentto transparent is that in a translucent or transparent condition, thecells in the device can be observed through the cover both during andafter loading of cells. This not only assists in the loading of cells,but also makes monitoring of the cells during use much easier.Additionally, position of various elements of the containment device aremore visible during assembly when the membrane is translucent.

[0072] In addition to using cells suspended in aqueous fluids in thepresent invention, cells suspended in viscous fluids, such as alginate,can also be loaded into the invention. With these cell suspensions, muchless fluid weeps through the permeable membrane as the suspension isloaded in the device. This often requires the cell suspension to be moreprecisely characterized in terms of cells numbers than with the aqueousfluids described above. Islets of Langerhans are examples of cell typesthat often benefit from being suspended in a viscous fluid when used inthe present invention.

[0073] The permeable membrane of the present invention should preventcells from moving into or out or the device, but allow the passage ofnutrients, waste products, and bioactive substances secreted by cellscontained within the device. In one embodiment, the membrane excludesparticles on a molecular scale. Such molecular weight cut off (MWCO)properties may be useful for excluding proteins, etc., produced by theimmune system of a recipient from traversing the membrane that wouldadversely effect cells encapsulated in the device. The precise MWCOrange appropriate for a particular application will vary depending onthe membrane material, type of cells contained within the device, thesize of the therapeutic cell product to be released into the surroundingenvironment, and the host environment, etc. Accordingly, selectivelypermeable membranes having a MWCO of between about 10 kD to about 2000kD may be suitable for use in the present invention. A MWCO range ofbetween about 30 kD and 150 kD is particularly preferred in applicationswhere it is desired to isolate the contained cells from contact withmolecules of the immune system capable of recognizing or destroying thecontained cells.

[0074] Referring now to FIG. 5, a preferred method of making acontainment device is as follows. This description is particularlyappropriate for a device that will carry a cellular suspension. Acentral core 204 is placed within a tubular membrane 202. The core is abiologically inert material, such as silicone, and has a cross-sectionalshape that is appropriate for the cell suspension. Examples of thecross-sectional shapes that are appropriate for a cell suspension areprovided in FIG. 4. In a preferred embodiment, the core has a stellarcross-section 401. However, the cross-sectional shape of the core is notcritical to the instant invention and may be any of a number ofdifferent shapes, such as those shown in FIG. 4. The core 204 length isonly slightly shorter that the length of a finished containment device.The tubular membrane 202 is longer than the core 204 to allow closure ofthe membrane at one end and attachment of a loading device at theopposite end of the membrane. In this embodiment, the core is notattached to the membrane in the final device. In other embodiments, thecentral core is attached to the membrane in the closure region.

[0075] Referring to FIG. 6, after core 204 is placed within the tubularmembrane 202, a polymer plug 506 is placed in one end of the tubularmembrane, abutting core 204. In a preferred embodiment, plug 506 isfluorinated ethylene propylene (FEP). The core, plug and membrane arearranged so that approximately 2 mm of membrane 202 extends beyond theFEP plug and the FEP plug abuts the silicone core. In this arrangement,a heat source or implement 602 is applied to the membrane in thevicinity of the FEP plug. The temperature of heat source 602 istypically between 350° C. and 450° C., preferably 390° C., which isabove the melt point of the FEP and causes the FEP to melt and flow. Thetemperature of the heat source is also slightly above the melt point ofthe ePTFE membrane. However, the heat exposure is not so great to causeany significant degradation or damage to the ePTFE membrane. As a resultthe membrane combines with the FEP plug to form a closure or seal 520 atthe distal end of the containment device. For purposes of distinguishingdifferent seals of the containment device, this seal will be termed adry seal. In one embodiment, the opening in heat source 602 is slightlysmaller than the diameter of FEP plug 506. In this embodiment, when heatsource 602 is applied, a slight clamping pressure helps to form dry seal520. After cooling, this closure or seal 520 is substantially orcompletely impermeable to cells that are eventually loaded in thedevice. The closure is also generally impermeable to liquids within thedevice. This closure is considered to be a dry seal, because it iscreated by a dry seal technique. In this context, the closure techniqueis dry because the ePTFE membrane, the silicone core in the vicinity ofthe closure region, and the FEP plug are dry when the closure is created(i.e., aqueous fluids are absent during the formation of the closure).

[0076] Referring to FIGS. 5 and 7, after forming the dry closure on oneend of the containment device, a blunt tip, such as an 18 gage needle508 about 25 mm long is inserted into a tube 510 of a coloredthermoplastic material. In a preferred embodiment, the tube is a coloredthermoplastic material made of FEP about 25 mm long. Needle 508 and FEPtube 510 assembly are inserted into the proximal end of the tubularePTFE membrane 202 that is opposite the dry seal 520.

[0077] The needle 508 and FEP 510 tube are advanced within the ePTFEmembrane 202 until there is a slight gap of about 2 mm between the endof the FEP tube 510 and the silicone core 204. This slight gap isimportant for the subsequent wet loading and sealing of the containmentdevice. At the same time, the silicone core 204 abuts the closure 520 atthe other end. In this arrangement, heat source 602 is applied to theend of the membrane 202 with a slight clamping pressure where it bondsor seals the membrane to the FEP tube 510 and also bonds or seals theFEP tube to the needle 508. The colored FEP becomes visible through themembrane after removing the heat and allowing the seal to cool. Thishelps to visually confirm a cell impermeable seal. Referring to FIG. 7,a seal 720 is formed when the ePTFE membrane, FEP tube, and needle aredry. Therefore, it is formed by a dry seal technique and is a dry seal.

[0078]FIGS. 7A-7C illustrate additional ways in which a thermoplasticpolymer material 512 can be placed in closure region 514 for wetsealing. Cell delivery means 515 are also provided in the figures.

[0079] Referring to FIG. 8, the same heat device used to form closure520 at the distal end of the containment device can be used to createseal 720. The heat device 602 is an electrically heated clamp or forcepswith a cylindrical opening 802 between the clamping jaws to surround thecontainment device 100. The jaws are about 2-3 mm thick. This designallows application of heat around the circumference of the containmentdevice and formation of a cell impermeable seal or closure. The heatdevice need not be electrically heated. For example, an ultrasonicheating device or a radio frequency induction device can be used togenerate heat in the desired location and thereby melt or fuse the FEPthermoplastic.

[0080] When used in combination with a containment apparatus of the typedisclosed by Butler et al. in U.S. Pat. No. 5,843,069, it is oftendesirable to form the sealed ends of the device into a smooth regularshape, such as a hemisphere. Applying a heated mold having the desiredshape to the sealed end of the present invention is a preferred way ofreshaping the sealed end. Heat can be generated in the mold withinfrared energy, ultrasound, or radio frequencies.

[0081] The method of making the containment device of the instantinvention has been described with reference to the figures. Referring toFIG. 9, that method is summarized as follows: At step 902, ePTFEmembrane material, made in the manner described above, is applied to asilver-plated copper (SPC) mandrel that is approximately 1.5-2.5 mm indiameter, and approximately 810 mm long. The first wrap is alongitudinal wrap of 0.5 inch wide ePTFE, and provides longitudinalstrength to the resulting tubular ePTFE membrane. The longitudinal wrapthat is applied at step 902 is overlapped at the tape edges to form thetubular shape.

[0082] At step 904, subsequent bias or helical wraps of ePTFE areapplied over the longitudinal ePTFE wrap. In a preferred embodiment,these bias wraps have about 50% overlap of the ePTFE tape. Multiple biaswraps are applied to the mandrel, with the wrap directions alternatingfor each successive layer. In a preferred embodiment, six (6)alternating layers of 0.5 inch wide ePTFE tape are applied to themandrel. At step 906, the ePTFE wrapped mandrel is placed into an ovenat approximately 385° C. for approximately eight (8) minutes. This heatstep allows the ePTFE wrap layers to bond, forming a tubular porousmembrane. The temperature and time are selected so that the resultingbond is of sufficient strength, without loss of the desired porosity. Atstep 908, the tubular ePTFE membrane is treated with a wetting agent,such as polyvinyl alcohol. This treatment helps to ensure that thenormally hydrophobic ePTFE will easily wet and form a porous permeablemembrane for the transfer of soluble materials.

[0083] At step 910, the tubular ePTFE membrane and mandrel are removedfrom the oven and cooled. The ends of the SPC mandrel are clamped, andthe mandrel is stretched approximately 30%. The stretch of the mandrelnecks down and slightly reduces the mandrel diameter. This reduction indiameter of the mandrel allows easy removal of the ePTFE tubularmembrane from the mandrel.

[0084] At step 912, the ePTFE tubular membrane is cut or trimmed to thedesired length. In one embodiment, the wrapped membrane is about 810 mmlong, which is sufficient to produce four (4) individual containmentdevices, with an 180 mm long membrane.

[0085] At step 914, a silicone core is inserted into the trimmed ePTFEtubular membrane. The core diameter is less than the diameter of thetubular membrane, allowing the core to easily slide into the membrane.In a preferred embodiment, the core is approximately 160 mm long and theePTFE membrane is approximately 180 mm long.

[0086] At step 916, an FEP plug, about 2 mm long, and also smaller indiameter than the ePTFE tubular membrane, is put into the distal end ofthe ePTFE tubular membrane. The FEP plug, silicone core and ePTFEmembrane are adjusted so that only a slight length (1-2 mm) of ePTFEmembrane extend beyond the FEP plug and the silicone core is close to,or abutts the FEP plug.

[0087] At step 918, the properly adjusted, or configured plug, core andmembrane are heat sealed at the distal end. The heat seal isaccomplished with the previously described electrically heated forceps.After heat seal, the distal end has an appearance similar to closure520, illustrated in FIG. 7.

[0088] At step 920, an 18 gage blunt tip needle is inserted into acolored FEP tube. The inner diameter of the FEP tube is slightly lessthan the outer diameter of the needle, to provide a snug sliding fit.

[0089] At step 922, the combined needle and FEP tube are inserted intothe proximal end of the ePTFE tubular membrane. The FEP tube is advancedinto the membrane until it is about 2 mm from the end of the siliconecore. This slight gap is helpful during cell loading to allow the cellsuspension to flow around the silicone core.

[0090] At step 924, the properly oriented ePTFE membrane, FEP tube andneedle are heat-sealed using the same electrically heated forceps toform seal 720. This is a secondary seal. After step 924, the containmentdevice is complete and ready for sterilization and subsequent loading.

[0091] This completes the method of making the containment device. Aftercompleted, the containment device can be checked for leaks and closureintegrity with any known type of leak detection, including a bubblepoint check. In a bubble point check, a containment device is lightlypressurized to determine if there is any leakage. An inadequate seal isrevealed by the evolution of bubbles from the sealed regions of thedevice. The completed containment device can also be sterilized by anumber of known techniques, including but not limited to chemicalsterilization and steam autoclave. Steam autoclave has an advantage ofwetting the membrane and displacing air within the containment devicewith sterile liquid.

[0092] Method of Loading the Device

[0093] A containment device 100, manufactured according to theabove-described method, is preferably loaded with cell suspensions ordrug formulations. As described above, an important aspect of theinstant invention is the cell-impermeable nature of the device. Thisaspect helps to ensure that any implant cells within the containmentdevice remain within the device. Assurance that there is no implant cellleakage from the containment device is important because the cells maybe genetically engineered or not compatible with the host immune system.If the cells in the suspension can reproduce, it may be desirable tolimit that reproduction to the interior of the containment device. Forthis reason, the cell-impermeable nature of the containment device,including the membrane and closures, is important.

[0094] The sealing mechanisms of the present invention help to ensurethat contamination from implanted cells does not occur from a faultyseal in a cell containment device. In addition, the present inventionalso helps to ensure that contaminating implant cells do not originatefrom the loading system or process. This is accomplished by eliminatingany open paths for the implant cells from a loading device to theexterior of the containment device during loading. This reduces oreliminates the possibility that implant cells will become attached tothe containment device exterior. Elimination or reduction in overallpossibility of contamination also prevents a possibility that theimplant cells can contaminate the loading area. This helps to ensurethat implant cells are not inadvertently transferred from the loadingarea to the exterior of the containment device during routine handling.

[0095] In the present invention, one method to accomplish theseobjectives is to use a closed cell-tight loading system. If the implantcell loading system remains closed, there is little or no possibilitythat implant cells can escape from the system and contaminate theloading area or exterior of the containment device.

[0096]FIG. 10 illustrates an embodiment for manually loading containmentdevice 100. It is understood that alternatively, the loading steps canbe performed in an automated facility. When the containment device willcontain a cell suspension, the empty sterilized containment device 100,immersed in sterile liquid 1002 is placed in a loading hood 1004. Theequipment and tools to load and seal the containment device are also inthe loading hood. Prior to placing the device in the loading hood, airis removed from the containment device and replaced by the sterileliquid. When the device is sterilized in liquid by steam sterilization,evacuation or removal of air in the device is a natural consequence ofthe sterilization. For other sterilization techniques that do notnecessarily remove air within the containment device, an air evacuationstep is generally desired.

[0097] As illustrated in FIG. 11, the containment device 100 is placedin a cell-loading fixture 1102, which is part of a loading jig 1104.This provides a stable platform for subsequent operations.

[0098] Referring to FIG. 12, within the loading hood, a loading device1202 is connected to the needle 508 of the containment device 100, suchas by a luer lock. In the manual system, loading device 1202 is a 1 mlsyringe. In an automated system, which is not illustrated, loadingdevice 1202 is part of an automated cell suspension handling anddelivery system. The connection between loading device 1202 and needle508 is cell-tight. Sterile water may be present in the device and theneedle and may spill onto the device at the time the two components areconnected. Generally speaking, it is important that cells are notintroduced into the immediate vicinity of the open needle hub 508 untilthe connection between the two components is first made. When connected,devices 100 and 1202 become a closed cell-tight system. In the manualsystem illustrated in FIG. 12, cells in suspension are extracted with apipette 1204 from a cell transfer container 1206.

[0099] Referring to FIG. 13, the pipette 1206 with cell suspension isused to place the cell suspension into the loading device 1202. It isimportant that cells are not allowed to leak or escape from pipette 1206as the cell suspension is transferred. A plunger 1302 is placed into theloading device 1202, thereby closing the loading system and forming aclosed cell-tight system.

[0100] Alternatively, the loading device can be charged with cells at adifferent station and then transported to the device loading station.Nevertheless, the cell delivery portion of the loading station issterile at the point of contact with the containment device. Theconnection between the loading device and the containment device iscell-tight to ensure a closed system.

[0101] A manual loading system is illustrated in FIGS. 10-13, to clearlyshow the various steps for attaching the containment device 100 to theloading device 1202 and then placing a cell suspension into the loadingdevice. However, using an open syringe, there is a remote possibility ofcontamination in the loading area if cells are inadvertently spilledduring the pipette transfer from the cell transfer device to the loadingdevice. Therefore, a preferred embodiment is a fully closed system,where the loading device includes suitable interlocks and valves toavoid even this remote possibility of contamination. Only after thecontainment device and loading device are connected, is there anyloading of cells into the containment device. There is no path for cellsto escape from the closed cell-tight system during loading. The onlypath for the cells is from the loading device to the interior of thecontainment device.

[0102] Alternatively, a process that permits cells to leak or escapeduring the loading process may be acceptable, provided the leakage iscontained and isolated from the cell containment device.

[0103] Referring to FIG. 14, after the cell suspension is within theloading device 1202, the cell suspension is loaded into containmentdevice 100 by a slight pressure. This pressure is supplied by manuallydepressing plunger 1302. As the cell suspension passes into thecontainment device, the suspension is concentrated. This concentrationis a consequence of the porous nature of the treated ePTFE tubularmembrane in containment device 100. Cells are unable to pass through themembrane, but the suspension fluid is able to pass through the membrane.This serves as a sieving action by the ePTFE membrane and retains orfilters the cells within the containment device, while allowing excesscell suspension fluid 1402 to pass through or weep from the ePTFEmembrane of containment device 100.

[0104] Alternatively, as illustrated in FIG. 21, if membrane 2102 doesnot readily permit liquid passage, end 2101 of the device is configuredto act as a receptacle for receiving a fluid stream. Permeable membrane2106 permits passage of liquids therethrough, while preventing cellsfrom escaping the device. After the containment device is loaded, theend is then sealed 2108. This is termed a “primary seal” because theclosure is formed after loading the device, while liquids of the cellsuspension are contacting the membrane and sealing polymer.

[0105] Methods of Sealing the Device

[0106] Once the cell suspension is loaded into the containment device, aclamp 1404 is applied to the containment device. Preferably, the clampis applied to a region of the tubular membrane containing the siliconecore such that the very end of the core is captured by the clampingforce. The clamp serves two purposes. One purpose is to serve as a heatsink during subsequent sealing operations. The other purpose is toprovide a method to hold the containment device during the sealingoperations.

[0107] After the cells are loaded, the system remains a closedcell-tight system during closure or wet-sealing of the containmentdevice. Referring to FIGS. 15 and 16, heat source 604 is applied with aslight pressure to a portion of the ePTFE membrane 202 in thecontainment device 100 that is in communication with the FEP polymertube 510. During heating, the device is simultaneously twisted andelongated to form a closure region 1602. Ideally, the twist is about 360degrees, but any twist of greater than about 45 degrees helps toaccomplish the objective of constricting and compressing the closure.

[0108] The twisting and elongation serves to provide a visibleseparation region. The elongation also ensures that when the containmentdevice is separated from the needle and loading device, a seal orclosure remains on each side of the separation. This wet-seal in closureregion 1602 is termed a primary seal. The heating serves to cauterizethe closure and kill any cells that may be within closure region 1602.The elongation and twisting also provides a slight pressure in the areaof the closure and serves to provide a visible separation region. Theelongation also ensures that when the containment device is separatedfrom the needle and loading device, a seal or closure remains on eachside of the separation, thereby ensuring that the system remains aclosed cell-tight system; cell leakage is prevented from the closure. Ina preferred embodiment, this elongation and closure is about 2 mm long,which can be visibly observed and readily bisected.

[0109] After the closure is created by heating, elongation, andtwisting, the closure is allowed to cool. At this point, the containmentdevice with the loaded cells or drugs remains attached to the needle bythe closure. However, the closure eliminates any fluid passage betweenthe needle and the containment device. This can be verified by slightlypressurizing the needle side of the closure and ensuring that noadditional weep emerges from the containment device. With an integralclosure, the containment device can be separated from the needle,without any fear of cell leakage from either the containment device, orthe loading device. Any cells within the closure region were eitherkilled outright by the heat closure, or rendered metabolicallynon-functional. The only step remaining in the seal or closure step isto separate the containment device from the needle.

[0110] Referring to FIG. 17, after the closure is formed, thecontainment device 100 is separated from the needle elements 1702.Separation is performed with scissors or a knife. Care is taken to makethe separation in the middle of the closure region, thereby maintainingcell-tight seals after the separation. The resulting closure oncontainment device 100 and needle elements 1702 include a fused orwelded area with FEP 510 and ePTFE 202.

[0111] The subsequent processing includes disposal of needle elements1702 and other associated elements of the loading device.

[0112] If the closure of the containment device extends, it may beappropriate to trim excess material from the closure. This is importantif the excess material can contact the containment apparatus duringplacement or replacement of the containment device.

[0113] The primary seal just described is formed as a wet seal, becauseit is formed in a containment device that has been loaded with a cellsuspension, resulting in a wet membrane. Even after the containmentdevice is sealed or closed and separated or cut from the loading device,the system remains closed. The closure substantially or completely sealsthe closure region and the heat used to create the closure helps toensure that any cells within the closure region are either killed, orhave their metabolic capabilities destroyed. This ensures that cells arenot released into the loading area or sterile field, and also helpsensure that multiple cell loads can be performed without risk ofcontamination from previous loading operations.

[0114] In the described embodiment, heat from an electrically heatedclamp is used to form the closures. In other embodiments, the closure ofthe containment device is created by other forms of heat application,such as ultrasonic welding or radio frequency inductive heating. Theonly requirement for heat closure is that the heat be applied locally inthe closure region, and that it melt the thermoplastic polymer to allowformation of the closure, without compromising the integrity of theePTFE membrane.

[0115] In another embodiment, the closure of the containment device iscreated by heating only the membrane material, without anythermoplastic. The only requirement is that the resulting closureprovide the required degree of closure or cell-impermeability. Inanother embodiment, a closure is formed by heating only a thermoplasticmaterial previously attached to the permeable membrane material. Asshown in FIG. 19C, the closure is made with a thermoplastic materialhaving a portion is attached to the permeable membrane and a portionthat extends beyond the permeable membrane to provide a port made solelyof thermoplastic material.

[0116] In another embodiment, the closure at the end of the containmentdevice that is normally created by a dry seal (i.e., the distal end)before the device is loaded, is created by a wet seal technique. In thisembodiment, the membrane is wet when the closure at the distal end iscreated. The source of the wet membrane is not limiting and may be theresult of a cell suspension load, a drug load, a wetting agent or asterile solution.

[0117] The closure can also be created by non-heat methods, such assolvents or chemicals. In this embodiment, an important aspect of theclosure is that after the closure is formed, there is little or nopossibility that living or viable cells remaining within the closureregion. If the solvent or chemical is toxic to the cells, then formationof the closure itself may be sufficient to ensure that the closureregion is free of viable cells. However, if the solvent or chemical isnot toxic to the cells, then an additional step must be provided toensure that any cells within the closure region are killed or renderednon-metabolically functioning. For example, it may be appropriate to usean ultraviolet light cured or activated compound or glue to create theclosure. These compounds or glue may not be toxic by themselves, but theUV cure may be sufficient to render the closure region cell dead ormetabolically functioning.

[0118] The method of filling and sealing the containment device of theinstant invention has been described above with reference to thefigures. Referring to FIG. 18, the steps of filling and sealing aresummarized. At step 1802, the ePTFE tubular membrane and needlecontainment device is sterilized. As indicated above, this is with anyof a number of different techniques, though steam sterilization ispreferred as initially wetting the membrane of the device.

[0119] At step 1804, air is removed from the containment device and themembrane is wet, if not accomplished in the sterilization step.

[0120] At step 1806, the containment device is attached to the celldelivery apparatus.

[0121] At step 1807, a cell suspension is transferred to the celldelivery apparatus.

[0122] At step 1808, a closed cell-tight system is formed.

[0123] At step 1809, a cell suspension is infused into the containmentdevice from the cell delivery apparatus. During this step, the cellsuspension can be concentrated, as the membrane serves as a sieve toallow excess suspension fluid to weep from the containment devicemembrane.

[0124] At step 1810, after the cell suspension is infused, a heat sinkis applied to the containment device. The heat sink is positioned justdistal to the eventual closure region.

[0125] At step 1812, the FEP tube is moved within the membrane so as toabutt the silicone core. As discussed above, there is a gap between thesilicone core and the FEP tube to assist with cell loading. Prior tosealing the containment device, this gap is preferably closed at step1812.

[0126] At step 1814, a heat source is applied to the ePTFE tubularmembrane in the vicinity of the closure region. The heat source is anyof a number of different types, with an electrically heated forceps apreferred embodiment.

[0127] At step 1816, while the heat source is maintained near theclosure region, the ePTFE membrane is both elongated and twisted to formthe closure. This combination of actions applies pressure in thevicinity of the closure and helps to ensure a good seal. The heat,elongation and twisting provides a cell kill zone. It is possible thatthe ePTFE membrane is only twisted, or only elongated to create theclosure. However, a combination provides the best closure.

[0128] At step 1818, the membrane closure is allowed to cool afterremoving the heat source. As the FEP and ePTFE cools, it hardens to formthe closure.

[0129] At step 1820, the containment device is separated from the celldelivery apparatus at the closure region. This is accomplished bycutting at the mid-point of the closure with a scissors or knife.

[0130] At step 1822, integrity of the seal at the cell deliveryapparatus is checked, such as by slightly pressurizing the apparatus andobserving any leaks.

[0131] At step 1824, ends of the containment device are trimmed toremove any irregularity or sharp features which might be problems duringsubsequent implant.

[0132] At this point, the containment device has been loaded with a cellsuspension, and the device has been sealed to form a cell-impermeableregion at the closure regions. The only steps remaining are preparationfor implant and implantation of the containment device directly into arecipient or indirectly in an implantable containment apparatus.

[0133] In the previous description, one embodiment and configuration ofthe containment device has been used to illustrate the inventiveaspects. FIG. 19 illustrates another embodiment of the instantinvention. In FIG. 19A, the tubular membrane 202, FEP tube 510 andneedle 508 are configured as generally described above. When thecontainment device is made, heat is applied to the ePTFE membrane,causing the FEP and membrane to fuse or melt and thereby create a seal1902 with needle 508. In another embodiment, illustrated in FIGS. 19Band 19C, the membrane is sealed to the FEP tube. However, the membranedoes not extend into the closure region and the closure is created withonly FEP. The FEP may be further connected in a cell-tight fashion, toother cell delivery components (see FIG. 19C). Cell-tight methods ofconnection include heat welds, luer locking, etc.

[0134] In an embodiment illustrated in FIG. 19C, the tubular membrane202 is heat sealed to FEP tube 510, to form seal 1906. However, membrane202 is not directly sealed to needle 508. In the embodiment illustratedin FIG. 19C, FEP tube 510 is heat sealed to needle 508 to form secondaryseal 1904. As illustrated in FIG. 19C, an additional seal is made duringcontainment device manufacture. In the subsequent loading of thecontainment device, it is clear that a closure in seal region 1906 willinclude both ePTFE and FEP. However, a closure in seal region 1908 willinclude only FEP. Depending on a number of factors, it may be desireableto use the embodiment illustrated in FIG. 19A for some applications, andthe embodiments illustrated in FIGS. 19B and 19C for other applications.

[0135]FIG. 20 illustrates an alternative embodiment where a dry seal2004 is formed on the end of a string or “sausage-link” of multiplecontainment devices 2006. The devices are simultaneously loaded andindividual closures 2004 are formed between devices 2006. The individualdevices are then separated at the closures 2004 between the devices2006. This has an advantage of volume processing.

[0136]FIG. 21 illustrates an alternative embodiment, which isparticularly advantageous where membrane weeping is not desired orpossible. In this embodiment, membrane 2102 surrounds core 2100. An FEPplug 2103 is between core 2100 and terminal 2105. Terminal 2105 isconnected to membrane 2102 and is either a filter 2106, to allowconcentration of a cell suspension as described above, or a fluidreceptacle (not illustrated) to capture cell or drug suspension afterflow through the containment device. A wet seal in effected in closureregion 2014.

[0137] After loading with the drug or cell suspension, a closure 2108 isformed in one of the manners described above for other embodiments. Inthis manner, the containment device can be readily loaded even whenmembrane 2102 does not or cannot weep.

[0138] It is also apparent, though not illustrated, that the tubularmembrane need not be outside the FEP tube. For example, the needle mightbe inserted into the tubular membrane and the FEP tube placed over boththe membrane and needle.

[0139] Clinical success with any implantable device containing somaticcells, engineered somatic cells, or immortalized transformed cells mustcontain those transplanted cells for the life of the device. Many of theproposed populations of cells to be used for somatic cell therapy in theinstant invention are both motile and immortal. Keeping those migratorycell populations within the device is a critical design parameter.

[0140] In vitro load testing was conducted in order to rigorouslymeasure the ability to load containment devices of the present inventionwithout any external cell contamination. Possible sources of cellcontamination on external surfaces of the devices include cells beingultra-filtrated through the ePTFE membranes, back washed out theproximal end of the device before the wet seal is done, pinholes, orsimply erroneous external contamination of the devices during the loadprocedure. Prokaryotic cells were used in the tests rather thanmammalian cells because of their much smaller size and much fastergrowth rates. Use of prokaryotic cells is believed to be a valid andextremely sensitive measure of external contamination from any sourceduring loading of the device.

[0141] Cell containment devices having wet seals were made as describedabove and presented with a bacterial challenge by loading broth culturescontaining either of two types of bacteria, M. luteus or P. aeruginosa,into the devices. These organisms were chosen due to size, shape andmotility differences that might affect their ability to beultrafiltrated and/or migrate through the ePTFE membranes of the presentinvention. Also, the colony growth and broth culture characteristicsallow easy and quick identification of each type of organism.

[0142] During loading, the fluid that normally ultra-filters through thepermeable membrane of a containment device of the present invention wasaseptically collected drop by drop onto standard microbiologic cultureplates of tryptic soy agar (TSA). The bottoms of the plates were markedindicating the exact location of where each drop fell. Once the deviceswere filled with the particular bacteria and the ultrafiltrate samplescollected and marked, the devices were wet sealed and placed into brothculture at 37 degree's centigrade along with their respective cultureplates (ultrafiltrate samples). The ultrafiltrate collected from theloading of the test devices were negative for growth of both organismstested. No colonies were present on the agar plates in the regionsmarked to indicate where the ultrafiltrate drops landed on the agar.

[0143] When cell-permissive devices (i.e., intentionally made leaky)were loaded with either M. luteus or P. aeruginosa and the ultrafiltratecollected onto sterile agar plates, there was rapid colony growth inexactly the spots where the drops of ultrafiltrate landed. The colonycolor (M. luteus is yellow) and morphology was consistent with theoriginal two organisms used and not a contaminant. In addition, thecolonies were sampled and sent out for identification. The externalresults confirmed the identifications as Pseudomonas spp. or Micrococcusspp.

[0144] Further studies were conducted by placing test devices andcontrol devices in an in vitro culture. Test devices for this study wereintentionally made to leak bacteria. Control devices were made accordingto the teachings of the present invention and were not made to leakbacteria. Following loading of the devices with one of the two strainsof bacteria, the test and control devices were placed into a tryptic soybroth (TSB) for culture. If a wet sealed containment device is notcell-tight, bacteria escape the device and bloom into a turbid culturewithin a matter of hours. The TSB media around the test devices becameturbid within a matter of hours for both species of bacteria tested. Incontrast, the culture media in which the control devices were culturedshowed no turbidity.

[0145] Although illustrative embodiments have been described herein indetail, it should be noted and will be appreciated by those skilled inthe art that numerous variations may be made within the scope of thisinvention without departing from the principle of this invention andwithout sacrificing its chief advantages.

[0146] Unless otherwise specifically stated, the terms and expressionshave been used herein as terms of description and not terms oflimitation. There is no intention to use the terms or expressions toexclude any equivalents of features shown and described or portionsthereof and this invention should be defined in accordance with theclaims that follow.

We claim:
 1. A cell containment device comprising: a permeable membranedelimiting a space for containing cells therein; a closure regioncomprising a thermoplastic polymer in association with the permeablemembrane and means for placing cells in said space through said closureregion; wherein said closure region is closeable with a wet seal.
 2. Acontainment device comprising: a membrane; a polymer in communicationwith the membrane; and at least one closure, the closure created byapplying heat to a portion of the membrane and to a portion of thepolymer after wetting the membrane with a liquid.
 3. A device accordingto claim 2, further comprising a cell suspension within a space enclosedby the membrane.
 4. A device according to claim 2, further comprising adrug formulation within a space enclosed by the membrane.
 5. A deviceaccording to claim 2, wherein the membrane includes a polymer membrane.6. A device according to claim 2, wherein the membrane includes a porousPTFE membrane.
 7. A device according to claim 2, wherein the membranesubstantially surrounds the polymer.
 8. A device according to claim 2,wherein the membrane is substantially tubular.
 9. A device according toclaim 2, further comprising a closure on a first end of the tubularmembrane and the closure created by applying heat is on a second end ofthe tubular membrane.
 10. A device according to claim 2, wherein thepolymer includes FEP.
 11. A device according to claim 2, wherein thepolymer is distinct from the membrane.
 12. A device according to claim2, wherein the closure is substantially part of the polymer.
 13. Adevice according to claim 2, wherein the closure is substantiallyseparate from the membrane.